Drug composition containing nucleic acid copolymer

ABSTRACT

This invention has for its object to insure an effective utilization of single-stranded nucleic acid copolymers, particularly poly(adenylic acid-uridylic acid), and to provide a pharmaceutical composition having antitumor activity. 
     The invention typically relates to a pharmaceutical composition comprising a lipid device such as Lipofectin (trademark), 3-O-(4-dimethylaminobutanoyl)-1,2-O-dioleylgycerol, 3-O-(2-dimethylamino-ethyl)carbamoyl-1,2-O-dioleylglycerol, 3-O-(2-diethylaminoethyl) carbamoyl-1,2-O-dioleylgycerol, or 2-O-(2-diethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol and poly(adenylic acid-uridylic acid).

This application is a 35 U.S.C. 371 application of PCT/JPG4/00238 filed17 Feb. 1994 published as WO94/18987 Sep. 1, 1994.

TECHNICAL FIELD

The present invention relates to a pharmaceutical compositioncharacterized by comprising a lipid device and a single-stranded nucleicacid copolymer.

The term lipid device as used herein means a device comprising a naturalor artificial lipid as a component thereof and having the function ofpromoting the intracellular uptake of a physiologically activesubstance.

BACKGROUND ART

Unlike a nucleic acid polymer-nucleic acid polymer complex such aspolyinosinic acid-polycytidylic acid, a single-stranded nucleic acidcopolymer such as poly (adenylic acid-uridylic acid) does not displayinterferon-inducing activity and hence, has no antitumor effect whenadministered alone.

Meanwhile, it is known that certain kinds of positively charged lipiddevices (e.g. cationic liposomes) are instrumental to the delivery ofgenes into cells (e.g. JP-A-4108391, WO91/17424). It is also known thatwhen a certain kind of nucleic acid such as a double-stranded RNA isadministered together with a lipid device such as cationic liposomes, apotentiated interferon inducer action is realized (U.S. Pat. No.5,049,386). It is generally conjectured that since the nucleic acids of,for example, genes are negatively charged, they form complexes withcationic liposomes and the complexes become fused to the cell membraneand the nucleic acids of genes or the like find their way into the cell.

However, it remains to be known whether application of a lipid device toa single-stranded nucleic acid copolymer gives rise to interferoninducer activity or an antitumor action.

DISCLOSURE OF INVENTION

The object of the present invention is to implement an effectiveutilization of a single-stranded nucleic acid copolymer and provide apharmaceutical composition having antitumor activity.

In the course of intensive research, the inventors of the presentinvention discovered that applying a lipid device to a single-strandednucleic acid copolymer such as poly(adenylic acid-uridylic acid) resultsin high interferon inducer activity and have accordingly developed theinstant invention.

The lipid device that can be used includes Lipofectin (trademark,manufactured by Bethesda Research Laboratories Life Technologies Inc.)and Genetransfer (trademark, manufactured by Wako Pure ChemicalIndustries), which are known, and a mixture of a compound of thefollowing general formula I! and a phospholipid. ##STR1## wherein R¹ andR² are not the same and each represents OY or --A--(CH₂)n-E. nrepresents a whole number of 0-4. E represents pyrrolidino, piperidino,substituted or unsubstituted piperazino, morpholino, substituted orunsubstituted guanidino, or ##STR2## (R³ and R⁴ are the same ordifferent and each represents hydrogen, lower(C₁₋₄) alkyl,hydroxy-lower(C₁ -₄) alkyl, or mono- or di-(lower) alkylaminoalkyl(C₂-₆)). A represents the followings 1, 2, 3, 4, 5 6 or 7. ##STR3##

R and Y are the same or different and each represents a saturated orunsaturated aliphatic hydrocarbon group of 10-30 carbon atoms or asaturated or unsaturated fatty acid residue of 10-30 carbon atoms.

The substituted piperazino for E includes 4-methylpiperazino,4-ethylpiperazino, 4-n-propylpiperazino, 4-isopropylpiperazino,4-n-butylpiperazino, 4-isobutylpiperazino, 4-(2-hydroxyethyl)piperazino,4-(2-hydroxypropyl)piperazino, and 4-(3-hydroxypropyl)piperazino, amongothers.

The substituted guanidino for E includes methylguanidino,ethylguanidino, n-propylguanidino, N,N-dimethylguanidino,N,N-diethylguanidino, N,N-di-n-propylguanidino, N,N'-dimethylguanidino,N,N'-diethylguanidino, N,N'-di-n-propylguanidino,N,N,N'-trimethylguanidino, N,N,N'-triethylguanidino,N,N,N'-tri-n-propylguanidino, N,N,N',N'-tetramethylguanidino,N,N,N',N'-tetraethylguanidino, and N,N,N',N'-tetra-n-propylguanidino,among others.

The lower alkyl for R³ and R⁴ includes methyl, ethyl, n-propyl,isopropyl, n-butyl, isobutyl, sec-butyl, and tert-butyl, among others.

The hydroxy(lower)alkyl for and R³ and R⁴ includes hydroxymethyl,1-hydroxyethyl, 2-hydroxyethyl, 1-hydroxypropyl, 2-hydroxypropyl, and3-hydroxypropyl, among others.

The mono- or di-(lower)alkylaminoalkyl for R³ and R⁴ includesmethylaminomethyl, dimethylaminomethyl, 2-(methylamino)ethyl,2-dimethylaminoethyl, 3-(methylamino)propyl, 3-dimethylaminopropyl,ethylaminomethyl, diethylaminomethyl, 2-(ethylamino)ethyl,2-diethylaminoethyl, 3-(ethylamino)propyl, 3-diethylaminopropyl,n-propylaminomethyl, di-n-propylaminomethyl, 2-(n-propylamino)ethyl,2-(di-n-propylamino)ethyl, 3-(n-propylamino)propyl, and3-(di-n-propylamino)propyl, among others.

Referring further to general formula I!, R and Y are the same ordifferent and each represents a saturated or unsaturated aliphatichydrocarbon group of 10-30 carbon atoms or a saturated or unsaturatedfatty acid residue of 10-30 carbon atoms as mentioned above. However,the preferred is the case in which R and Y are the same and eachrepresents an unsaturated aliphatic hydrocarbon or unsaturated fattyacid residue of about 12-20 carbon atoms. The most preferred is the casein which both R and Y represent oleyl or oleoyl, for instance.

A is preferably a carbamate bond or an ester bond.

Specifically, the following compounds can be mentioned as typicalexamples.

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dilaurylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dimyristylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dipalmitylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dilinoleylglycerole,

3-O-(2-Dimethylaminoethyl)carbamoyl-2-O-lauryl-1-O-myristylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1-O-oleyl-2-O-palmitylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1-O-linoleyl-2-O-oleylglycerol,

3-O-(Dimethylaminomethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(3-Dimethylaminopropyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(4-Dimethylaminobutyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Diethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(3-Diethylaminopropyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Di-n-propylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Diisopropylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Di-n-butylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Diisobutylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Di-sec-butylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O- 2-(N-Ethyl-N-methylamino)ethyl!carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Methylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Ethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-n-Propylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-n-Butylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Aminoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-2-(N-Methyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,2-O-dioleylglycerol,

3-O-2-(N-Ethyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,2-O-dioleylglycerol,

3-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,2-O-dioleylglycerol,

3-O-2-(N-(2-Diethylamino)ethyl-N-methylamino)ethyl!carbamoyl-1,2-O-dioleylglycerol,

3-O- 2-(4-Methylpiperazino)ethyl!carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Morpholinoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Piperidinoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Pyrrolidinoethyl)carbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Diethylaminoethyl)thiocarbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Dimethylaminoethyl)thiocarbamoyl-1,2-O-dioleylglycerol,

3-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!thiocarbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Pyrrolidinoethyl)thiocarbamoyl-1,2-O-dioleylglycerol,

3-O-(2-Dimethylaminoethyl)sulfamoyl-1,2-O-dioleylglycerol,

3-O-(2-Diethylaminoethyl)sulfamoyl-1,2-O-dioleylglycerol,

3-O-2-N,N-Di-(2-hydroxyethyl)amino)ethyl!sulfamoyl-1,2-O-dioleylglycerol,

3-O-(2-Pyrrolidinoethyl)sulfamoyl-1,2-O-dioleylglycerol,

3-O-(N,N-Dimethylaminoacetyl)-1,2-O-dioleylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1,2-O-dilaurylglycerol,

3-O-(4-Diethylaminobutanoyl)-1,2-O-dipalmitylglycerol,

3-O-(4-diemthylaminobutanoyl)-1,2-O-dioleylglycerol,

3-O-(4-Diethylaminobutanoyl)-1,2-O-dioleylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1,2-O-dilinoleylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1-O-oleyl-2-O-palmitylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1-O-linoleyl-2-O-oleylglycerol,

3-O-(3-Dimethylaminopropionyl)-1,2-O-dioleylglycerol,

3-O-(5-Dimethylaminopentanoyl)-1,2-O-dioleylglycerol,

3-O-(4-Di-n-propylaminobutanoyl)-1,2-O-dioleylglycerol,

3-O-(4-Diisopropylaminobutanoyl)-1,2-O-dioleylglycerol,

3-O- 4-(N-Ethyl-N-methylamino)butanoyl!-1,2-O-dioleylglycerol,

3-O- 4-Ethylaminobutanoyl)-1,2-O-dioleylglycerol,

3-O-4-(N-Methyl-N-(2-hydroxyethyl)amino)butanoyl!-1,2-O-dioleylglycerol,

3-O- 4-(N,N-Di-(2-hydroxyethyl)amino)butanoyl!-1,2-O-dioleylglycerol,

3-O-4-(N-(2-Diethylamino)ethyl-N-methylamino)butanoyl!-1,2-O-dioleylglycerol,

3-O- 4-(4-Methylpiperazino)butanoyl!-1,2-O-dioleylglycerol,

3-O-(4-Morpholinobutanoyl)-1,2-O-dioleylglycerol,

3-O-(4-Pyrrolidinobutanoyl)-1,2-O-dioleylglycerol,

3-O-(4-Piperidinobutanoyl)-1,2-O-dioleylglycerol,

O-(2-Diethylaminoethyl)-O'-(2,3-dioleyloxypropyl)methylphosphonate,

O-(2-Dimethylaminoethyl)-O'-(2,3-dioleyloxypropyl)methylphosphonate,

O-2-(N,N-di(2-hydroxyethyl)amino)ethyl!-O'-(2,3-dioleyloxypropyl)methylphosphonate,

O-(2-Pyrrolidinoethyl)-O'-(2,3-dioleyloxypropyl)methylphosphonate,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dilauroylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dimyristoylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dipalmitoylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1,2-O-dilinolenylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1-O-oleoyl-2-O-palmitoylglycerol,

3-O-(2-Dimethylaminoethyl)carbamoyl-1-O-linolenyl-2-O-oleoylglycerol,

3-O-(3-Dimethylaminopropyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Diethylaminoethyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Di-n-propylaminoethyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Diisopropylaminoethyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O- 2-(N-Ethyl-N-methylamino)ethyl!carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Ethylaminoethyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O-2-(N-Methyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,2-O-dioleoylglycerol,

3-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,2-O-dioleoylglycerol

3-O-2-(N-(2-Diethylamino)ethyl-N-methylamino)ethyl!carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Piperidinoethyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Pyrrolidinoethyl)carbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Aminoethyl)carbamoyl-1,2-O-dioleoylglycerol

3-O-(2-Diethylaminoethyl)thiocarbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Dimethylaminoethyl)thiocarbamoyl-1,2-O-dioleoylglycerol,

3-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!thiocarbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Pyrrolidinoethyl)thiocarbamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Diethylaminoethyl)sulfamoyl-1,2-O-dioleoylglycerol,

3-O-(2-Dimethylaminoethyl)sulfamoyl-1,2-O-dioleoylglycerol,

3-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!sulfamoyl-1,2-O-dioleoylglycerol

3-O-(2-Pyrrolidinoethyl)sulfamoyl-1,2-O-dioleoylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1,2-O-dilauroylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1,2-O-dimyristoylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1,2-O-dipalmitoylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1,2-O-dioleoylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1,2-O-dilinolenylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1-O-oleoyl-2-O-palmitoylglycerol,

3-O-(4-Dimethylaminobutanoyl)-1-O-linolenyl-2-O-oleoylglycerol,

3-O-(3-Dimethylaminopropionyl)-1,2-O-dioleoylglycerol,

3-O-(5-Dimethylaminopentanoyl)-1,2-O-dioleoylglycerol,

3-O-(4-Diethylaminobutanoyl)-1,2-O-dioleoylglycerol,

3-O-(4-Di-n-propylaminobutanoyl)-1,2-O-dioleoylglycerol,

3-O-(4-Diisopropylaminobutanoyl)-1,2-O-dioleoylglycerol,

3-O- 4-(N-Ethyl-N-methylamino)butanoyl!-1,2-O-dioleoylglycerol,

3-O-(4-Ethylaminobutanoyl)-1,2-O-dioleoylglycerol,

3-O-4-(N-Methyl-N-(2-hydroxyethyl)amino)butanoyl!-1,2-O-dioleoylglycerol,

3-O- 4-(N,N-Di-(2-hydroxyethyl)amino)butanoyl!-1,2-O-dioleoylglycerol,

3-O-4-(N-(2-Diethylamino)ethyl-N-methylamino)butanoyl!-1,2-O-dioleoylglycerol

3-O-(4-Pyrrolidinobutanoyl)-1,2-O-dioleoylglycerol,

O-(2-Diethylaminoethyl)-O'-(2,3-dioleoyloxypropyl)methylphosphonate,

O-(2-Dimethylaminoethyl)-O'-(2,3-dioleoyloxypropyl)methylphosphonate,

O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!-O'-(2,3-dioleoyloxypropyl)methylphosphonate,

O-(2-Pyrrolidinoethyl)-O'-(2,3-dioleoyloxypropyl)methylphosphonate,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dilaurylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dimyristylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dipalmitylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dilinoleylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1-O-lauryl-3-O-myristylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1-O-oleyl-3-O-palmitylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1-O-linoleyl-3-O-oleylglycerol,

2-O-(3-Dimethylaminopropyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(4-Dimethylaminobutyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Diethylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Di-n-propylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-diisopropylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Di-n-butylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Diisobutylaminoethyl)carbamoyl-,3-O-dioleylglycerol,

2-O-(2-Di-sec-butylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O- 2-(N-Ethyl-N-methyl)aminoethyl!carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Methylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Ethylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-n-Propylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Butylaminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Aminoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-2-(N-Methyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleylglycerol,

2-O-2-(N-Ethyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleylglycerol,

2-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleylglycerol,

2-O-2-(N-(2-Diethylamino)ethyl-N-methylamino)ethyl!carbamoyl-1,3-O-dioleylglycerol,

2-O- 2-(4-Methylpiperazino)ethyl!carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Morpholinoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Piperidinoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Pyrrolidinoethyl)carbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Dimethylaminoethyl)thiocarbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Diethylaminoethyl)thiocarbamoyl-1,3-O-dioleylglycerol,

2-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!thiocarbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Pyrrolidinoethyl)thiocarbamoyl-1,3-O-dioleylglycerol,

2-O-(2-Dimethylaminoethyl)sulfamoyl-1,3-O-dioleylglycerol,

2-O-(2-Diethylaminoethyl)sulfamoyl-1,3-O-dioleylglycerol,

2-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!sulfamoyl-1,3-O-dioleylglycerol,

2-O-(2-Pyrrolidinoethyl)sulfamoyl-1,3-O-dioleylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1,3-O-dilaurylglycerol,

2-O-(4-Diethylaminobutanoyl)-1,3-O-dipalmitylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1,3-O-dilinoleylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1-O-oleyl-3-O-palmitylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1-O-linoleyl-3-O-oleylglycerol,

2-O-(3-Dimethylaminopropionyl)-1,3-O-dioleylglycerol,

2-O-(5-Dimethylaminopentanoyl)-1,3-O-dioleylglycerol,

2-O-(4-Di-n-propylaminobutanoyl)-1,3-O-dioleylglycerol,

2-O-(4-Diisopropylaminobutanoyl)-1,3-O-dioleylglycerol,

2-O- 4-(N-Ethyl-N-methyl)aminobutanoyl!-1,3-O-dioleylglycerol,

2-O-(4-Ethylaminobutanoyl)-1,3-O-dioleylglycerol,

2-O-4-(N-Methyl-N-(2-hydroxyethyl)amino)butanoyl!-1,3-O-dioleylglycerol,

2-O- 4-(N,N-Di-(2-hydroxyethyl)amino)butanoyl!-1,3-O-dioleylglycerol,

2-O-4-(N-(2-Diethylamino)ethyl-N-methylamino)butanoyl!-1,3-O-dioleylglycerol,

2-O- 4-(4-Methylpiperazino)butanoyl!-1,3-O-dioleylglycerol,

2-O-(4-Morpholinobutanoyl)-1,3-O-dioleylglycerol,

2-O-(4-Pyrrolidinobutanoyl)-1,3-O-dioleylglycerol,

2-O-(4-Piperidinobutanoyl)-1,3-O-dioleylglycerol,

O-(2-Diethylaminoethyl)-O'-(1,3-dioleyloxypropyl)methylphosphonate,

O-(2-Dimethylaminoethyl)-O'-(1,3-dioleyloxypropan-2-yl)methylphosphonate,

O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!-O'-(1,3-dioleyloxypropan-2-yl)methylphosphonate,

O-(2-Pyrrolidinoethyl)-O'-(1,3-dioleyloxypropan-2-yl)methylphosphonate,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dilauroylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dimyristoylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dipalmitoylglycerol,

2-O-(2-Diethylaminoethyl)carbamoyl-1,3-O-dipalmitoylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1,3-O-dilinolenylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1-O-oleoyl-3-O-palmitoylglycerol,

2-O-(2-Dimethylaminoethyl)carbamoyl-1-O-linolenyl-3-O-oleoylglycerol,

2-O-(Dimethylaminomethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-(3-Dimethylaminopropyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Diethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Di-n-propylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Diisopropylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O- 2-(N-Ethyl-N-methylamino)ethyl!carbamoyl-1,3-O-dioleoylglycerol,

2-O- 2-(N-Methyl-N-n-butylamino)ethyl!carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Ethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

2-O-2-(N-Methyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleoylglycerol,

2-O-2-(N-Ethyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleoylglycerol,

2-O-2-(N-(2-Diethylamino)ethyl-N-methylamino)ethyl!carbamoyl-1,3-O-dioleoylglycerol,

2-O-2-(N,N,N',N'-Tetramethylguanidino)ethyl!carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Morpholinoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Piperidinoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Pyrrolidinoethyl)carbamoyl-1,3-O-dioleoylglycerol,

2-O- 2-(4-Ethylpiperazino)ethyl!carbamoyl-1,3-O-dioleoylglycerol,

2-O-2-(4-(2-Hydroxyethyl)piperazino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

2-O-(2-Diethylaminoethyl)thiocarbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Dimethylaminoethyl)thiocarbamoyl-1,3-O-dioleoylglycerol,

2-O-2-(N,N-Di-(2-hydroxyethyl)amino)ethyl!thiocarbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Pyrrolidinoethyl)thiocarbamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Diethylaminoethyl)sulfamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Dimethylaminoethyl)sulfamoyl-1,3-O-dioleoylglycerol,

2-O-2-N,N-Di-(2-hydroxyethyl)aminoethyl!sulfamoyl-1,3-O-dioleoylglycerol,

2-O-(2-Pyrrolidinoethyl)sulfamoyl-1,3-O-dioleoylglycerol,

2-O-(3-Diethylaminopropionyl)-1,3-O-dioleoylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1,3-O-dilauroylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1,3-O-dimyristoylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1,3-O-dipalmitoylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1,3-O-dioleoylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1,3-O-dilinolenylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1-O-oleoyl-3-O-palmitoylglycerol,

2-O-(4-Dimethylaminobutanoyl)-1-O-linolenyl-3-O-oleoylglycerol,

2-O-(3-Dimethylaminopropionyl)-1,3-O-dioleoylglycerol,

2-O-(5-Dimethylaminopentanoyl)-1,3-O-dioleoylglycerol,

2-O-(4-Diethylaminobutanoyl)-1,3-O-dioleoylglycerol,

2-O-(4-Di-n-propylaminobutanoyl)-1,3-O-dioleoylglycerol,

2-O-(4-Diisopropylaminobutanoyl)-1,3-O-dioleoylglycerol,

2-O- 4-(N-Ethyl-N-methylamino)butanoyl!-1,3-O-dioleoylglycerol,

2-O- 4-(Ethyl)aminobutanoyl!-1,3-O-dioleoylglycerol,

2-O-4-(N-Methyl-N-(2-hydroxyethyl)amino)butanoyl!-1,3-O-dioleoylglycerol,

2-O- 4-(N,N-Di-(2-hydroxyethyl)amino)butanoyl!-1,3-O-dioleoylglycerol,

2-O-4-(N-(2-Diethylamino)ethyl-N-methylamino)butanoyl!-1,3-O-dioleoylglycerol,

2-O-(4-Pyrrolidinobutanoyl)-1,3-O-dioleoylglycerol,

O-(2-Dimethylaminoethyl)-O'-(1,3-dioleoyloxypropan-2-yl)methylphosphonate,

O-(2-Aminoethyl)-O'-(1,3-dioleoyloxypropan-2-yl)methylphosphonate,

O-(2-Diethylaminoethyl)-O'-(1,3-dioleoyloxypropan-2-yl)methylphosphonate,

2-Dimethylaminoethyl N-(2,3-dilauryloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dimyristyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dilinoleyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2-lauryloxy-3-linoleyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(3-myristyloxy-2-oleyloxypropyl)carbamate,

3-Dimethylaminopropyl N-(2,3-dioleyloxypropyl)carbamate,

4-Dimethylaminobutyl N-(2,3-dioleyloxypropyl)carbamate,

2-Diethylaminoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Di-n-propylaminoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Di-n-butylaminoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Ethylmethylaminoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-(N-Ethyl-N-methylamino)ethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Ethylaminobutyl N-(2,3-dioleyloxypropyl)carbamate,

2-n-Propylaminoethyl N-(2,3-dioleyloxypropyl)carbamate,

2- N-Methyl-N-(2-hydroxyethyl)amino!ethylN-(2,3-dioleyloxypropyl)carbamate,

2- N-Ethyl-N-(2-hydroxyethyl)amino!ethylN-(2,3-dioleyloxypropyl)carbamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethyl N-(2,3-dioleyloxypropyl)carbamate,

2- N-(2-Diethylamino)ethyl-N-methylamino!ethylN-(2,3-dioleyloxypropyl)carbamate,

2-(4-Methylpiperadino)ethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Morpholinoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Piperidinoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Pyrrolidinoethyl N-(2,3-dioleyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dioleyloxypropyl)thiocarbamate,

2-Diethylaminoethyl N-(2,3-dioleyloxypropyl)thiocarbamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethylN-(2,3-dioleyloxypropyl)thiocarbamate,

2-Pyrrolidinoethyl N-(2,3-dioleyloxypropyl)thiocarbamate,

2-Dimethylaminoethyl N-(2,3-dioleyloxypropyl)sulfamate,

2-Diethylaminoethyl N-(2,3-dioleyloxypropyl)sulfamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethyl N-(2,3-dioleyloxypropyl)sulfamate,

2-Pyrrolidinoethyl N-(2,3-dioleyloxypropyl)sulfamate,

N-(2,3-Dioleyloxy)propyl-4-dimethylaminobutylamide,

N-(2,3'Dioleyloxy)propyl-4-diethylaminobutylamide,

N-(2,3'Dioleyloxy)propyl-4- N,N-di(2-hydroxyethyl)amino!butylamide,

N-(2,3'Dioleyloxy)propyl-4-pyrrolidinobutylamide,

2-Dimethylaminoethyl N-(2,3-dilauroyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dimyristoyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dipalmitoyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dioleoyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dilinolenyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2-oleoyloxy-3-palmitoyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2-linolenyloxy-3-oleoyloxypropyl)carbamate,

2-Diethylaminoethyl N-(2,3-dioleoyloxypropyl)carbamate,

3-Dimethylaminopropyl N-(2,3-dioleoyloxypropyl)carbamate,

2-Diisopropylaminoethyl N-(2,3-dioleoyloxypropyl)carbamate,

2-Di-n-propylaminoethyl N-(2,3-dioleoyloxypropyl)carbamate,

2-(N-Ethyl-N-methylamino)ethyl N-(2,3-dioleoyloxypropyl)carbamate,

2-Ethylaminoethyl N-(2,3-dioleoyloxypropyl)carbamate,

2- N-methyl-N-(2-hydroxyethyl)amino!ethylN-(2,3-dioleoyloxypropyl)carbamate,

2- N,N-Di-(2hydroxyethyl)amino!ethyl N-(2,3-dioleoyloxypropyl)carbamate,

2- N-(2-Diethylamino)ethyl-N-methylamino!ethylN-(2,3-dioleoyloxypropyl)carbamate,

2-Piperidinoethyl N-(2,3-dioleoyloxypropyl)carbamate,

2-Pyrrolidinoethyl N-(2,3-dioleoyloxypropyl)carbamate,

2-Aminoethyl N-(2,3-dioleoyloxypropyl)carbamate,

2-Dimethylaminoethyl N-(2,3-dioleoyloxypropyl)thiocarbamate,

2-Diethylaminoethyl N-(2,3-dioleoyloxypropyl)thiocarbamate,

2- N,N-Di-(2-hydroxyethyl)aminoethyl!N-(2,3-dioleoyloxypropyl)thiocarbamate,

2-Pyrrolidinoethyl N-(2,3-dioleoyloxypropyl)thiocarbamate,

2-Dimethylaminoethyl N-(2,3-dioleoyloxypropyl)sulfamate,

2-Diethylaminoethyl N-(2,3-dioleoyloxypropyl)sulfamate,

2- N,N-Di-(2-hydroxyethyl)aminoethyl!N-(2,3-dioleoyloxypropyl)sulfamate,

2-Pyrrolidinoethyl N-(2,3-dioleoyloxypropyl)sulfamate,.

N-(2,3-Dioleoyloxy)propyl-4-dimethylaminobutylamide,

N-(2,3-Dioleoyloxy)propyl-4-diethylaminobutylamide,

N-(2,3-Dioleoyloxy)propyl-4- N,N-di-(2-hydroxyethyl)amino!butylamide,

N-(2,3-Dioleoyloxy)propyl-4-pyrrolidinobutylamide,

2-Dimethylaminoethyl N-(1,3-dilauryloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dimyristyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dilinoleyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1-lauryloxy-3-linoleyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1-myristyloxy-3-oleyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1-oleyloxy-3-palmityloxypropan-2-yl)carbamate,

3-Dimethylaminopropyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

4-Dimethylaminobutyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Diethylaminoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Di-n-propylaminoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Di-n-butylaminoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-(N-Ethyl-N-methylamino)ethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Methylaminoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Ethylaminobutyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-n-Propylaminoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-n-Butylamino N-(1,3-dioleyloxypropan-2-yl)carbamate,

2- N-Methyl-N-(2-hydroxyethyl)amino!ethylN-(1,3-dioleyloxypropan-2-yl)carbamate,

2- N-Ethyl-N-(2-hydroxyethyl)amino!ethylN-(1,3-dioleyloxypropan-2-yl)carbamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethylN-(1,3-dioleyloxypropan-2-yl)carbamate,

2- N-(2-Diethylamino)ethyl-N-methylamino!ethylN-(1,3-dioleyloxypropan-2-yl)carbamate,

2-(4-Methylpiperadino)ethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Piperidinoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Pyrrolidinoethyl N-(1,3-dioleyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dioleyloxypropan-2-yl)thiocarbamate,

2-Diethylaminoethyl N-(1,3-dioleyloxypropan-2-yl)thiocarbamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethylN-(1,3-dioleyloxypropan-2-yl)thiocarbamate,

2-Pyrrolidinoethyl N-(1,3-dioleyloxypropan-2-yl)thiocarbamate,

2-Dimethylaminoethyl N-(1,3-dioleyloxypropan-2-yl)sulfamate,

2-Diethylaminoethyl N-(1,3-dioleyloxypropan-2-yl)sulfamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethylN-(1,3-dioleyloxypropan-2-yl)sulfamate,

2-Pyrrolidinoethyl N-(1,3-Dioleyloxypropan-2-yl)sulfamate,

N-(4-Dimethylaminobutanoyl)-1,3-dioleyloxy-1-amino-propane,

N-(4-Diethylaminobutanoyl)-1,3-dioleyloxy-1-amino-propane,

N-4-(N,N-Di-(2-hydroxyethyl)amino)butanoyl!-1,3-dioleyloxy-1-amino-propane

N-4-Pyrrolidinobutanoyl-1,3-dioleyloxy-1-amino-propane,

2-Dimethylaminoethyl N-(1,3-dilauroyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dimyristoyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dipalmitoyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dilinolenyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1-oleoyloxy-3-palmitoyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1-linolenyloxy-3-oleoyloxypropan-2-yl)carbamate,

2-Diethylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

3-Dimethylaminopropyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-Diisopropylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-Di-n-propylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-(N-Ethyl-N-methylamino)ethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-Ethylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2- N-Methyl-N-(2-hydroxyethyl)amino!ethylN-(1,3-dioleoyloxypropan-2-yl)carbamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethylN-(1,3-dioleoyloxypropan-2-yl)carbamate,

2- N-(2-Diethylamino)ethyl-N-methylamino!ethylN-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-Piperidinoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-Pyrrolidinoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate

2-Aminoethyl N-(1,3-dioleoyloxypropan-2-yl)carbamate,

2-Dimethylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)thiocarbamate,

2-Diethylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)thiocarbamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethylN-(1,3-dioleoyloxypropan-2-yl)thiocarbamate,

2-Pyrrolidinoethyl N-(1,3-dioleoyloxypropan-2-yl)thiocarbamate,

2-Dimethylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)sulfamate,

2-Diethylaminoethyl N-(1,3-dioleoyloxypropan-2-yl)sulfamate,

2- N,N-Di-(2-hydroxyethyl)amino!ethylN-(1,3-dioleoyloxypropan-2-yl)sulfamate,

2-Pyrrolidinoethyl N-(1,3-dioleoyloxypropan-2-yl)sulfamate,

N-(2,3-Dioleoyloxy)propyl-4-dimethylaminobutylamide,

N-(2,3-Dioleoyloxy)propyl-4-diethylaminobutylamide,

N-(2,3-Dioleoyloxy)propyl-4- N,N-di-(2-hydroxyethyl)amino!butylamide,

N-(2,3-Dioleoyloxy)propyl-4-pyrrolidinobutylamide.

Among compounds of general formula I!,3-O-(4-dimethylaminobutanoyl)-1,2-O-dioleylglycerol,3-O-(2-dimethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,3-O-(2-diethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol,2-O-(2-diethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol, etc. arepreferred. Particularly preferred is3-O-(2-diethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol.

The compound I! can be obtained by the following and other processes.

(1) Where R¹ represents OY and A represents --O--C(═O)--NH-- ##STR4##(wherein B is typically imidazolyl, halogen or phenoxy. The halogen mayfor example be chlorine, bromine or iodine. R, Y, E and n are as definedhereinbefore)

As shown schematically above, compound I! can be synthesized by reactingII! with III!.

This reaction between II! and III! can be carried out using 1-3equivalents of III! per equivalent of II! in the presence of a solventat 0° C.-150° C. for 1-20 hours. The reaction solvent that can be usedincludes dimethylformamide, pyridine, toluene, benzene, ether, dioxane,tetrahydrofuran, chloroform and so on. To hasten the reaction, a basesuch as triethylamine can be added. Moreover, III! may be firstconverted to a metal salt using sodium hydride, n-butyllithium or thelike in the above-mentioned solvent and, then, reacted with II!.

(2) Where R¹ represents OY and A represents --NH--C(═O)--O-- ##STR5##(wherein B, R, Y, E and n are as defined hereinbefore)

Compound I! can be synthesized by reacting IV! with V! according to theabove reaction schema, using reaction conditions similar to thosementioned under (1).

(3) Where R¹ represents OY and A represents --NH--C(═O)--O-- ##STR6##(wherein R, Y, E and n are as defined hereinbefore)

Compound I! can be synthesized by reacting VI! with III'! as shownschematically above, using reaction conditions similar to thosementioned under (1).

(4) Where R¹ represents OY and A represents --O--C(═O)--NH-- ##STR7##(wherein R, Y, E and n are as defined hereinbefore)

Compound I! can be synthesized by reacting IV'! with VII! as shownschematically above, using reaction conditions similar to thosementioned under (1).

(5) Where R¹ represents OY and R² represents --A--(CH₂)n-E ##STR8##(wherein A, E and n are as defined hereinbefore)

Compound I! can be synthesized by transforming the hydroxyl groups ofthe above compound into the substituent groups R and Y through reactionwith suitable acylating agents (e.g. the anhydrides or acid chlorides offatty acids). This route of synthesis is preferred where R and Y arefatty acid residues. <Synthesis of the starting compounds IV!, IV'!, V!,VI!, VII!, and VIII!>

(1) Synthesis of Starting Compound IV'!

The starting compound IV'! can be synthesized typically in accordancewith the following reaction schema. ##STR9## (wherein Tr representstrityl, Ts represents tosyl; R and Y are as defined hereinbefore)

(2) Synthesis of Starting Compound IV!

The starting compound IV! can be synthesized typically by the followingexemplary process.

Starting with compound IV'!. the compound IV! can be synthesized by theconventional procedure, e.g. azidation and subsequent reduction.

(3) Synthesis of Starting Compound V!

The starting compound V! wherein B is imidazolyl, for instance, can besynthesized by reacting compound III'! with N,N'-carbonyldiimidazole inpyridine at ambient temperature.

(4) Synthesis of Starting Compound VI!

The starting compound VI! can be synthesized by reacting compound IV!with diphosgene.

(5) Synthesis of Starting Compound VII!

The starting compound VII! can be easily synthesized typically byreacting compound III! with diphosgene or by reacting a compound of theformula HOOC--(CH₂)n-E (where n and E are as defined hereinbefore) withDPPA (diphenylphosphoryl azide) in the presence of a tertiary amine suchas triethylamine at 0°-150° C. and further in the presence of a tertiaryamine such as pyridine at 0°-150° C.

(6) Synthesis of Starting Compound VIII!

1 The compound VIII! wherein A represents --O--C(═O)--NH-- can besynthesized typically according to the following reaction schema.##STR10## (wherein Im represents imidazolyl)

2 The compound VIII! wherein A represents --NH--C(═O)--O-- can besynthesized typically according to the following reaction schema.##STR11## (wherein Im represents imidazolyl; E and n are as definedhereinbefore)

3 The compound VIII! wherein A represents --O--C(═O)-- can besynthesized typically according to the following reaction schema.##STR12## (wherein DCC means dicyclohexylcarbodiimide and DMAP means4-N,N-dimethylaminopyridine; E and n are as defined hereinbefore)

4 The compound VIII! wherein A represents --O--C(═S)--NH-- can besynthesized typically according to the following reaction schema.##STR13## (wherein Im represents imidazolyl)

5 The compound VIII! wherein A represents --NH--C(═O)-- can besynthesized typically according to the following reaction schema.##STR14## (wherein DCC means dicyclohexylcarbodiimide; E and n are asdefined hereinbefore)

6 The compound VIII! wherein A represents --OSO₂ --NH-- can besynthesized typically according to the following reaction schema.##STR15## (wherein E and n are as defined hereinbefore)

7 The compound VIII! wherein A represents --O--P(═O)(--CH₃)--O-- can besynthesized typically according to the following reaction schema.##STR16## (wherein BT means 1-benzotriazolyl)

The phospholipid for use in combination with compound I! may for examplebe phosphatidylethanolamine or phosphatidylcholine.

The constitutional ratio of compound I! to phospholipid is appropriately0.1:9.9-9.9:0.1 (compound I!: phospholipid (molar ratio)), preferably1:9-9:1 (compound I!: phospholipid (molar ratio)), and for still betterresults, 1:3-3:1 (compound I!: phospholipid (molar ratio)).

In the pharmaceutical composition according to the present invention(hereinafter referred to as composition of the invention), the lipiddevice may be any of lipid suspension, liposome and other forms.

The lipid device comprised of compound I! and phospholipid can be simplyprepared typically by admixing compound I! with phospholipid in thepresence of water. It can also be prepared by a process which comprisesdissolving compound I! and phospholipid in chloroform, removing thechloroform thoroughly under a blast of nitrogen gas, stirring themixture well with addition of water, and subjecting it to sonication forseveral minutes.

The single-stranded nucleic acid copolymer may for example bepoly(adenylic acid-uridylic acid), poly(inosinic acid-uridylic acid) orthe like. The sequence of two constitutional bases may be regular orirregular. The term "regular" means that the two constituent bases arearranged alternately or a block formed by a given number of units of onebase and that of the other base are alternate. Preferred is asingle-stranded nucleic acid copolymer in which the two constituentbases are arranged alternately. Still more preferred is poly(adenylicacid-uridylic acid) of such alternate structure. The number of bases ofthe single-stranded nucleic acid copolymer that can be used in thepractice of the present invention is not critically restricted but issuitably in the range of 10 to 5,000.

The single-stranded nucleic acid copolymer for use in accordance withthe present invention may have undergone coalescence to form an apparentdouble-strand. Moreover, the single-stranded nucleic acid copolymer mayhave undergone partial coalescence to form a local double-strand.

The ratio of the lipid device to the single-stranded nucleic acidcopolymer is preferably 1:0.1-1:10 (lipid device: single-strandednucleic acid copolymer) by weight.

The composition of the invention can be manufactured by adding thesingle-stranded nucleic acid copolymer to the lipid device and stirringthe mixture by suitable means. The composition may also be provided byadding the single-stranded nucleic acid copolymer in the course ofpreparation of the lipid device.

The composition of the invention is preferably administered in singledosage forms and can be applied to animals including man by theintravenous, intraarterial, oral, intratissue, local (e.g. transdermal)or rectal route. Particularly preferred are intravenous administration,intraarterial administration, and local administration. Of course, thecomposition is administered in dosage forms suitable for the respectiveroutes, such as injections, peroral preparations, inhalants, eyedrops,ointments, suppositories and so on.

While the dosage of the composition of the invention is preferablydetermined in consideration of the species of active ingredient, dosageform, patient factors such as age and body weight, route ofadministration, nature and severity of disease, etc., the usual dosagefor adults is generally 0.1 mg-10 g/day/man, preferably 1 mg-500mg/day/man in terms of the active ingredient. A lower dosage may besufficient in some cases, while a higher dosage may be needed in others.The dosage may be administered in a few divided doses or at intervals ofa few days.

BEST MODE OF PRACTICING THE INVENTION

The following examples are intended to illustrate the present inventionin further detail.

Reference Example 1 Synthesis of 1,2-O-dioleylglycerol

(1) In 50 ml of pyridine was dissolved 4.6 g (50 mmol) of glycerolfollowed by addition of 13.9 g (50 mmol) of trityl chloride and themixture was stirred at ambient temperature overnight. The reactionmixture was then concentrated under reduced pressure and the residue wasdiluted with water and extracted with ether. The organic layer waswashed with water, dried over magnesium sulfate, and concentrated. Theresidue was purified by column chromatography (silicagel/chloroform-methanol) to provide 9.5 g (59%) of 1-O-tritylglycerol.

(2) In 120 ml of xylene was dissolved 3.22 g (10 mmol) of1-O-tritylglycerol, followed by addition of 3.36 g (30 mmol) oft-butoxypotassium under argon. After 5 minutes of stirring, 30 ml of asolution of 12.8 g (30 mmol) oleyl p-toluenesulfonate in xylene wasadded dropwise and the mixture was stirred under reduced pressure (20-30mmHg) at ambient temperature for 30 minutes and, then, at 50 for 1 hour.The reaction mixture was then poured in ice-water and extracted withether, and the extract was washed with water, dried, and concentrated.The residue was purified by column chromatography (silicagel/n-hexane-ethyl acetate) to provide 6.10 g (73%) of1,2-O-dioleyl-3-O-tritylglycerol.

(3) 1,2-O-Dioleyl-3-O-tritylglycerol (6.10 g, 7.3 mmol) was reacted with5% trichloroacetic acid/methylene chloride (50 ml, w/v) at ambienttemperature for 1 hour. The organic layer was then washed with saturatedaqueous sodium hydrogen carbonate solution and water, dried, andconcentrated. The residue was purified by column chromatography (silicagel/chloroform) to provide 3.75 g (87%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz, CH₃ ×2), 1.14-1.44 (44H,m, CH₂ ×22), 1.48-1.68 (4H, m, OCH₂ CH₂ ×2), 1.90-2.10 (8H, m, CH═CHCH₂×4), 3.38-3.78 (9H, m, OCH₂ ×4 & OCH), 5.26-5.45 (4H, m, CH═CH×2) MS(FAB): 593 (M+H)⁺

Reference Example 2 Synthesis of 2,3-dioleyloxypropylamine

(1) To a mixture of 1.00 g (1.7 mmol) of 1,2-O-dioleylglycerol, 0.83 g(17 mmol) of lithium azide, 0.89 g (3.4 mmol) of triphenylphosphine, and1.13 g (3.4 mmol) of carbon tetrabromide was added 10 ml ofN,N-dimethylformamide in bolus and the mixture was stirred at ambienttemperature for 3 hours. After completion of the reaction, the solventwas distilled off and the residue was diluted with water and extractedwith ether. The ether layer was washed with water, dried, andconcentrated, and the residue was purified by column chromatography(silica gel/n-hexane-ethyl acetate) to provide 1.03 g (100%) of2,3-dioleyloxypropyl azide as oil.

IR(neat, cm⁻¹): 2920, 2850, 2100

(2) In 30 ml of tetrahydrofuran was suspended 75 mg (2 mmol) of lithiumaluminum hydride. While this suspension was held under ice-cooling, 1.03g (1.7 mmol) of 2,3-dioleyloxypropyl azide was added dropwise and themixture was stirred for 30 minutes. Then, the mixture was furtherstirred at ambient temperature for 2 hours. At completion of thereaction, the reaction mixture was poured in ice-water and extractedwith ether and the extract was washed with water, dried, andconcentrated. The residue was subjected to column chromatography (silicagel/methylene chloride-methanol) to provide 0.98 g (98%) of the titlecompound as colorless oil.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz, CH₃ ×2), 1.17-1.45 (44H,m, CH₂ ×22), 1.48-1.70 (4H, m, OCH₂ CH₂ ×2), 1.90-2.14 (8H, m, CH═CHCH₂×4), 2.64-2.91 (2H, m, NCH₂), 3.30-3.78 (9H, m, OCH₂ ×3 & OCH),5.25-5.46 (4H, m, CH═CH×2) MS (FAB): 592 (M+H)⁺

Reference Example 3 Synthesis of 1,3-O-dioleylglycerol

(1) In pyridine were dissolved 1.00 g (11 mmol) of glycerol and 2.96 g(43 mmol) of imidazole and azeotropic distillation was carried out. Theresidue obtained was dissolved in 15 ml of N,N-dimethylformamide. Tothis solution under ice-cooling was added 3.60 g (24 mmol) oftributyldimethylsilyl chloride, and the mixture was stirred at ambienttemperature for 5 hours. After completion of the reaction, the solventwas distilled off and the residue was diluted with methylene chlorideand washed with saturated aqueous sodium hydrogen carbonate solution.This was dried and-concentrated to provide 3.45 g (99%) of1,3-O-di-(t-butyldimethylsilyl)glycerol.

(2) In dioxane was dissolved 3.45 g (11 mmol) of1,3-O-di-(t-butyldimethylsilyl)glycerol followed by addition of 3.03 g(12 mmol) of pyridinium p-toluenesulfonate. To this suspension was added16.5 ml (22 mmol) of dihydrofuran gradually under ice-cooling and themixture was stirred for 1 hour. After return to ambient temperature, themixture was allowed to react overnight. After completion of thereaction, the solvent was distilled off and the residue was treated withmethylene chloride and saturated aqueous sodium hydrogen carbonatesolution. The methylene chloride layer was washed with water, dried, andconcentrated to provide 4.25 g (100%) of1,3-O-di-(t-butyldimethylsilyl)-2-O-tetrahydrofuranylglycerol.

(3) To a solution of 4.25 g (11 mmol)1,3-O-di-(t-butyldimethylsilyl)-2-O-tetrahydrofuranylglycerol in 30 mltetrahydrofuran was added 30 ml of tetra-n-butylammonium fluoride (1mol/l in THF) dropwise and the mixture was stirred at ambienttemperature for 2 hours. The reaction mixture was then concentrated andthe residue was subjected to column chromatography (silica gel/methylenechloride-methanol) to provide 1.70 g (96%) of2-O-tetrahydrofuranylglycerol.

(4) In 30 ml of xylene was dissolved 854 mg (5.3 mmol) of2-O-tetrahydrofuranylglycerol. To this was added 1.78 g (15.9 mmol) oft-butoxypotassium under argon gas and the mixture was stirred for 5minutes. Then, 10 ml of a solution of 6.71 g (15.9 mmol) oleylp-toluenesulfonate in xylene was added dropwise and the mixture wasstirred under reduced pressure (20-30 mmHg) at ambient temperature for30 minutes and further at 50° C. for 1 hour. This reaction mixture waspoured in ice-water and extracted with ether, and the extract was washedwith water, dried, and concentrated. The residue was purified by columnchromatography (silica gel/chloroform) to provide 628 mg (18%) of1,3-O-dioleyl-2-O-tetrahydrofuranylglycerol as yellow oil.

(5) In 30 ml of tetrahydrofuran was dissolved 628 mg (0.95 mmol) of1,3-O-dioleyl-2-O-tetrahydrofuranylglycerol followed by addition of 5 mlof diluted (10%) hydrochloric acid, and the mixture was stirredovernight. The reaction mixture was then diluted with water, neutralizedwith saturated aqueous sodium hydrogen carbonate solution, and extractedwith ether. The extract was dried and concentrated and the residue wassubjected to column chromatography (silica gel/n-hexane-ethyl acetate)to provide 321 mg (57%) of the title compound as colorless oil.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz, CH₃ ×2), 1.14-1.26 (44H,m, CH₂ ×22), 1.49-1.68 (4H, m, OCH₂ CH₂ ×2), 1.98-2.13 (8H, m, CH═CHCH₂×4), 3.37-3.56 (8H, m, OCH₂ ×4), 3.95 (1H, brs, OC H), 5.27-5.46 (4H, m,CH═CH×2) MS (FAB): 593 (M+H)⁺

Reference Example 4 Synthesis of 1,3-dioleyloxy-2-propylamine

(1) In 5 ml of pyridine was dissolved 150 mg (0.25 mmol) of1,3-O-dioleylglycerol followed by addition of 77 mg (0.40 mmol) ofp-toluenesulfonyl chloride and the mixture was heated at 60° C. andstirred for 2 days. After completion of the reaction, the solvent wasdistilled off and the residue was diluted with water and extracted withether. The extract was dried and concentrated to provide 150 mg (80%) of1,3-O-dioleyl-2-O-(p-toluenesulfonyl)glycerol as yellow oil.

(2) A mixture of 150 mg (80%) of the above1,3-O-dioleyl-2-O-(p-toluenesulfonyl)glycerol, 30 mg (0.6 mmol) oflithium azide, and 5 ml of N,N-dimethylformamide was stirred at 100° C.for 2 hours. After cooling, the solvent was distilled off and theresidue was diluted with water and extracted with ether. The extract waswashed with water, dried, and concentrated to provide 125 mg (99%) of1,3-dioleyloxy-2-propyl azide as light-brown oil.

(3) In 3 ml of tetrahydrofuran was suspended 8 mg (0.2 mmol) of lithiumaluminum hydride. While this suspension was maintained underice-cooling, 125 mg (0.2 mmol) of 1,3-dioleyloxy-2-propyl azide wasadded dropwise and the mixture was stirred at 0° C. for 2 hours. Thereaction mixture was then poured in ice-water and extracted with ether.The extract was washed with water, dried, and concentrated. The residuewas subjected to column chromatography (silica gel/methylenechloride-methanol) to provide 104 mg (89%) of the title compound ascolorless oil.

Reference Example 5 Synthesis of 1,2-O-dioleoylglycerol

(1) In pyridine was dissolved 1 g (0.011 mol) of glycerin and azeotropicdistillation was carried out. The residue was dissolved in 30 ml ofpyridine, followed by addition of 4.05 g (0.012 mol) of dimethoxytritylchloride under ice-cooling. The mixture was then stirred at ambienttemperature overnight. After completion of the reaction, 5 ml ofmethanol was added and the mixture was stirred for 30 minutes, at theend of which time the solvent was distilled off. To the residue wasadded methylene chloride and the mixture was washed with saturatedaqueous sodium hydrogen carbonate solution, dried, and concentrated. Theresidue was subjected to column chromatography (silica gel/methylenechloride-methanol, 0.1% pyridine) to provide 2.58 g (60.2%) of1-O-dimethoxytritylglycerol.

(2) The 1-O-dimethoxytritylglycerol thus obtained, 290 mg (0.735 mmolwas subjected to azeotropic distillation with pyridine and the residuewas dissolved in 5 ml of pyridine. Then, 669 mg (2.223 mmol) of oleoylchloride was added with ice-cooling and the reaction was carried out at50° C. for 6 hours. After completion of the reaction, the solvent wasdistilled off under reduced pressure and the residue was diluted withmethylene chloride, washed with saturated aqueous sodium hydrogencarbonate solution, dried, and concentrated. The residue was subjectedto column chromatography (silica gel/n-hexane-methylene chloride) toprovide 519 mg (76.5%) of 1-O-dimethoxytrityl-2,3-O-dioleoylglycerol.

¹ H-NMR (60 MHz, CDCl₃) δ:0.88 (6H, m), 1.27 (40H, brs.), 1.50-1.70 (4H,m), 1.90-2.10 (8H, m), 3.10-3.30 (2H, m), 3.79 (6H, s), 4.20-4.40 (2H,m), 5.10-5.50 (5H, m), 6.70-7.40 (13H, m)

(3) The above 1-O-dimethoxytrityl-2,3-O-dioleoylglycerol, 218 mg (0.236mmol), was dissolved in 10 ml of 5% formic acid-methylene chloride andthe reaction was conducted for 10 minutes. The reaction mixture was thenneutralized with saturated aqueous sodium hydrogen carbonate solutionand the organic layer was further washed with saturated aqueous sodiumhydrogen carbonate solution, dried, and concentrated. The residue wassubjected to column chromatography (silica gel/n-hexane-methylenechloride-methanol) to provide 100 mg (68.0%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.28-2.40 (4H, m), 3.72 (2H, d,J=6 Hz), 4.10-4.40 (2H, m), 5.00-5.12 (1H, m), 5.30-5.40 (4H, m) MS(FAB): 621 (M+H)⁺

Reference Example 6 Synthesis of 1,3-O-dioleoylglycerol

(1) In 60 ml of pyridine was dissolved 2.75 g (0.013 mol) of2-O-(t-butyldimethylsilyl)glycerol, followed by addition of 8.82 g(0.028 mol) of oleoyl chloride under ice-cooling. The reaction wasconducted at 50° C. for 15 hours. After completion of the reaction, thesolvent was distilled off and the residue was diluted with methylenechloride, washed with saturated aqueous sodium hydrogen carbonatesolution, dried, and concentrated. The above procedure provided1,3-O-dioleoyl-2-O-(t-butyldimethylsilyl)glycerol.

(2) To 1,3-O-dioleoyl-2-O-(t-butyldimethylsilyl)glycerol was added 266ml of 0.1M tetra-n-butylammonium fluoride-tetrahydrofuran and thereaction was carried out at ambient temperature for 30 minutes. Aftercompletion of the reaction, the solvent was distilled off under reducedpressure and the residue was diluted with methylene chloride, washedwith water, dried, and concentrated. The residue was subjected to columnchromatography (silica gel/ethyl acetate-n-hexane) to provide 3.97 g(48.0% based on 2-O-t-butyldimethylsilylglycerol) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.34 (4H, t, J=8 Hz), 4.10-4.22(5H, m), 5.30-5.40 (4H, m) MS (FAB): 621 (M+H)⁺

Reference Example 7 Synthesis of1,3-O-dioleoyl-2-O-(2-bromoethyl)carbamoylglycerol

(1) In pyridine was dissolved 230 mg (0.37 mmol) of1,3-O-dioleoylglycerol and the solution was subjected to azeotropicdistillation. The residue was dissolved in 5 ml of pyridine and after120 mg (0.740 mmol) of N,N'-carbonyldiimidazole was added, the mixturewas stirred at ambient temperature for 3 hours. The solvent was thendistilled off under reduced pressure and the residue was dissolved inmethylene chloride, washed with 5% sodium dihydrogen phosphate-water,dried, and concentrated. The residue was dissolved in 10 ml ofN,N-dimethylformamide and after 45 mg (0.737 mmol) of 2-aminoethanol wasadded, the mixture was stirred at ambient temperature overnight. Aftercompletion of the reaction, the solvent was distilled off and theresidue was dissolved in methylene chloride, washed with 5% sodiumdihydrogen phosphate, dried, and concentrated. The residue was subjectedto column chromatography (silica gel/methylene chloride-methanol) toprovide 204 mg (79.5%) of1,3-O-dioleoyl-2-O-(2-hydroxyethyl)carbamoylglycerol.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (40H, brs.),1.50-1.80 (4H, m), 1.90-2.10 (8H, m), 2.34 (4H, t, J=8 Hz), 3.28-3.40(2H, m), 3.64-3.80 (2H, m), 4.20-4.40 (4H, m), 5.06-5.20 (2H, m),5.30-5.50 (4H, m) MS (FAB): 690 (M--OH)⁺

(2) To a mixture of 160 mg (0.226 mmol)1,3-O-dioleoyl-2-O-(2-hydroxyethyl)carbamoylglycerol, 150 mg (0.452mmol) of carbon tetrachloride, and 120 mg (0.458 mmol) oftriphenylphosphine was added 10 ml of N,N-dimethylformamide in bolus andthe mixture was stirred at ambient temperature for 2 hours. Aftercompletion of the reaction, the solvent was distilled off and theresidue was dissolved in methylene chloride, washed with water, dried,and concentrated. The residue was subjected to column chromatography(silica gel/ethyl acetate-n-hexane) to provide 91 mg (52.2%) of thetitle compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.86 (6H, t, J=6 Hz), 1.28 (4OH, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.31 (4H, t, J=8 Hz), 3.40-3.52(2H, m), 3.52-3.70 (2H, m), 4.20-4.44 (4H, m), 5.06-5.20 (2H, m),5.25-5.40 (4H, m) MS (FAB): 770 (M+H)

Example of Synthesis-1 Synthesis of3-O-(2-dimethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol

To 25 ml of a solution of 2.00 g (3.4 mmol) of 1,2-O-dioleylglycerol inpyridine was added 0.66 g (4.1 mmol) of N,N'-carbonyldiimidazole and themixture was stirred at ambient temperature for 5 hours. The solvent wasthen distilled off under reduced pressure and the residue was dissolvedin methylene chloride, washed with 5% sodium dihydrogen phosphate-water,dried, and concentrated. The residue was dissolved in 20 ml ofN,N-dimethylformamide, and after addition of 595 mg (6.8 mmol) ofN,N-dimethylethylenediamine, the mixture was stirred overnight. Aftercompletion of the reaction, the solvent was distilled off and theresidue was diluted with water and extracted with methylene chloride.The extract was washed with water, dried, and concentrated and theresidue was subjected to column chromatography (silicagel/chloroform-methanol) to provide 2.18 g (91%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=7 Hz, CH₃ ×2), 1.16-1.44 (44H,m, CH₂ ×22), 1.47-1.68 (4H, m, OCH₂ CH₂ ×2), 1.84-2.12 (8H, m, CH═CHCH₂×4), 2.20 (6H, s, N(CH₃)₂), 2.39 (2H, t, J=6 Hz, NCH₂), 3.18-3.31 (2H,m, CONHC H₂), 3.36-3.64 (7H, m, OCH₂ ×3 & OCH), 4.03-4.26 (2H, m, CH₂OCO), 5.22 (1H, brs., NHCO), 5.28-5.43 (4H, m, CH═CH×2) MS (FAB): 707(M+H)⁺

Example of Synthesis-2 Synthesis of 3-O-(2-methylaminoethyl)carbamoyl-1,2-O-dioleylglycerol

The title compound was obtained in the same manner as Example ofSynthesis-1 except that N-methylethylenediamine was used in lieu ofN,N-dimethylethylenediamine.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (44H, brs.),1.50-1.60 (4H, m), 1.90-2.10 (8H, m), 2.43 (3H, s), 2.71 (2H, t, J=6Hz), 3.28 (2H, q, J=6 Hz), 3.40-3.70 (7H, m), 4.05-4.26 (2H, m), 5.14(1H, brs.), 5.30-5.44 (4H, m) MS (FAB): 693 (M+H)⁺

Example of Synthesis-3 Synthesis of3-O-(2-aminoethyl)carbamoyl-1,2-O-dioleylglycerol

The compound synthesized using N-tritylethylenediamine in lieu ofN,N-dimethylethylenediamine in otherwise the same manner as Example ofSynthesis-1 was treated with 5% trichloroacetic acid-methylene chlorideand purified in the same manner to provide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (44H, brs.),1.50-1.60 (4H, m), 1.90-2.10 (8H, m), 3.10-3.20 (2H, m), 3.40-3.70 (9H,m), 4.04-4.26 (2H, m), 5.30-5.45 (4H, m), 6.20 (1H, brs.) MS (FAB): 679(M+H)⁺

Example of Synthesis-4 Synthesis of3-O-(2-diethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol

Using N,N-diethylethylenediamine in lieu of N,N-dimethylethylenediamine,the procedure of Example of Synthesis-1 was otherwise repeated toprovide the title compound.

¹ H-NMR (200 MHz, CDCl₃ m) δ:0.87 (6H, t, J=6 Hz), 1.01 (6H, t, J=6 Hz),1.27 (44H, brs.), 1.46-1.62 (4H, m), 1.90-2.10 (8H, m), 2.48-2.62 (6H,m), 3.18-3.30 (2H, m), 3.38-3.66 (7H, m), 4.04-4.24 (2H, m), 5.24-5.44(5H, m) MS (FAB): 735 (M+H)⁺

Example of Synthesis-5 Synthesis of3-O-(4-dimethylaminobutyl)carbamoyl-1,2-O-dioleylglycerol

Using 4-dimethylaminobutylamine in lieu of N,N-dimethylethylenediamine,the procedure of Example of Synthesis-1 was otherwise repeated toprovide the title compound. ¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6Hz), 1.28 (44H, brs.), 1.46-1.70 (8H, m), 1.90-2.10 (8H, m), 2.39 (6H,s), 2.44-2.56 (2H, m), 3.10-3.24 (2H, m), 3.36-3.70 (7H, m), 4.00-4.24(2H, m), 5.18-5.42 (5H, m) MS (FAB): 736 (M+H)⁺

Example of Synthesis-6 Synthesis of3-O-(2-dimethylaminoethyl)thiocarbamoyl-1,2-O-dioleylglycerol

Using N,N'-thiocarbonyldiimidazole in lieu of N,N'-carbonyldiimidazole,the procedure of Example of Synthesis-1 was otherwise repeated toprovide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (44H, brs.),1.50-1.60 (4H, m), 1.90-2.10 (8H, m), 2.21 (6H, d, J=4 Hz), 2.36-2.54(2H, m), 3.30-3.80 (9H, m), 4.40-4.70 (2H, m), 5.26-5.45 (4H, m) MS(FAB): 723 (M+H)⁺

Example of Synthesis-7 Synthesis of3-O-(4-dimethylaminobutanoyl)-1,2-O-dioleylglycerol

In 6 ml of methylene chloride-N,N-dimethylformamide (1:2) was dissolved120 mg (0.20 mmol) of 1,2-O-dioleylglycerol, followed by addition of 168mg (1 mmol) of 4-dimethylaminobutyric acid hydrochloride. Then, 206 mg(1 mmol) of N,N'-dicyclohexylcarbodiimide (DCC) and 25 mg (0.2 mmol) of4-dimethylaminopyridine were further added and the reaction wasconducted at ambient temperature overnight. The precipitated byproducturea was filtered off using a glass filter and the filtrate wasconcentrated to dryness under reduced pressure and treated withmethylene chloride-saturated aqueous sodium hydrogen carbonate solution.After phase separation, the methylene chloride layer was dried oversodium sulfate and the solvent was distilled off under reduced pressure.The residue was subjected to column chromatography (silica gel/methylenechloride-methanol) to provide 123 mg (87%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.20-1.40 (44H, m),1.45-1.60 (4H, m), 1.70-1.90 (2H, m), 1.90-2.10 (8H, m), 2.22 (6H, s),2.30 (2H, t, J=8 Hz), 2.36 (2H, t, J=8 Hz), 3.38-3.85 (7H, m), 4.04-4.30(2H, m), 5.30-5.45 (4H, m) MS (FAB): 706 (M+H)⁺

Example of Synthesis-8 Synthesis of3-O-(N,N-dimethylaminoacetyl)-1,2-O-dioleylglycerol

In a solvent mixture of 22 ml N,N-dimethylformamide and 11 ml methylenechloride was suspended 572 mg (5.547 mmol) of N,N-dimethylglycinefollowed by addition of 1736 mg (8.414 mmol) ofN,N'-dicyclohexylcarbodiimide and the mixture was stirred at ambienttemperature overnight. The solvent was then distilled off under reducedpressure and the residue was dissolved in 12 ml of pyridine containing327 mg (0.551 mmol) of dissolved 1,2-O-dioleylglycerol. Then, 80 mg(0.388 mmol) of N,N'-dicyclohexylcarbodiimide (DCC) was added and thereaction was conducted at 50° C. overnight. After completion of thereaction, the solvent was distilled off and the residue was dissolved inmethylene chloride, washed with saturated aqueous sodium hydrogencarbonate solution, dried, and concentrated. The residue was subjectedto column chromatography (silica gel/ethyl acetate-n-hexane) to provide251 mg (67.2%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (44H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.36 (6H, s), 3.23 (2H, s),3.40-3.70 (7H, m), 4.00-4.20 (2H, m), 5.20-5.40 (4H, m) MS (FAB): 678(M+H)⁺

Example of Synthesis-9 Synthesis of3-O-(4-diethylaminobutanoyl)-1,2-O-dioleylglycerol

(1) In 5 ml of anhydrous pyridine was dissolved 300 mg (0.51 mmol) of1,2-O-dioleylglycerol, followed by addition of 188 mg (1.01 mmol) of4-bromobutyl chloride under ice-cooling. After the temperature wasallowed to return to ambient temperature, the reaction was conducted at50° C. for 1 hour. The solvent was then distilled off and the residuewas treated with methylene chloride-saturated aqueous sodium hydrogencarbonate solution. After phase separation and drying over sodiumsulfate, the solvent was distilled off under reduced pressure. Theresidue thus obtained was subjected to column chromatography (silicagel/methylene chloride-methanol) to provide 159 mg (42%) of the bromocompound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.27 (44H, brs.),1.50-1.70 (4H, m), 1.90-2.20 (10H, m), 2.53 (2H, t, J=8 Hz), 3.40-3.70(9H, m), 4.05-4.30 (2H, m), 5.25-5.45 (4H, m)

(2) In 6 ml of N,N-dimethylformamide-isopropyl alcohol-chloroform(1:1:1) was dissolved 130 mg (0.18 mmol) of the above bromo compoundfollowed by addition of 1 ml of diethylamine and 70 mg (0.54 mmol) ofN,N-diisopropylethylamine. The mixture was reacted at 60° C. for 20hours and at 80° C. for a further 6 hours. The solvent was thendistilled off under reduced pressure and the residue was treated withmethylene chloride-water. The methylene chloride layer was dried oversodium sulfate and concentrated under reduced pressure. The residue wassubjected to column chromatography (silica gel/methylenechloride-methanol) to provide 63 mg (50%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.04 (6H, t, J=6 Hz),1.27 (44H, brs.), 1.50-1.70 (4H, m), 1.80 (2H, m), 1.90-2.10 (8H, m),2.37 (2H, t, J=6 Hz), 2.44-2.70 (6H, m), 3.40-3.70 (7H, m), 4.05-4.30(2H, m), 5.30-5.45 (4H, m) MS (FAB): 734 (M+H)⁺

Example of Synthesis-10 Synthesis ofN-(2,3-dioleyloxy)propyl-4-dimethylaminobutylamide

In 3 ml of anhydrous N,N-dimethylformamide was dissolved 100 mg (0.17mmol) of 2,3-dioleyloxypropylamine. To this solution were added 71 mg(0.42 mmol) of 4-dimethylaminobutyric acid hydrochloride, 105 mg (0.51mmol) of N,N'-dicyclohexylcarbodiimide (DCC), and 4.1 mg (0.034 mmol) of4-dimethylaminopyridine and the reaction was conducted at ambienttemperature overnight. The reaction mixture was then treated as inExample of Synthesis-6 to provide 115 mg (96%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.20-1.40 (44H, m),1.50-1.60 (4H, m), 1.70-1.90 (2H, m), 1.90-2.10 (8H, m), 2.23 (6H, s),2.24 (2H, t, J=8 Hz), 2.34 (2H, t, J=8 Hz), 3.20-3.60 (9H, m), 5.30-5.42(4H, m) MS (FAB): 705 (M+H)⁺

Example of Synthesis-11 Synthesis of3-O-(2-dimethylaminoethyl)sulfamoyl-1,2-O-dioleylglycerol

In 4 ml of methylene chloride-pyridine (2:1) was dissolved 150 mg (0.25mmol) of 1,2-O-dioleylglycerol. Then, 1 ml of a solution of 150 mg (0.75mmol) (2-dimethylaminoethyl)sulfamoyl chloride in methylene chloride wasadded and the reaction was carried out at ambient temperature for 2hours. After completion of the reaction, the solvent was distilled offunder reduced pressure and the residue was treated with methylenechloride-saturated aqueous sodium hydrogen carbonate solution. Themethylene chloride layer was dried over sodium sulfate and concentratedunder reduced pressure. The residue was subjected to columnchromatography (silica gel/methylene chloride-methanol) to provide 34 mg(18%) of the title compound. ¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6Hz), 1.20-1.40 (44H, m), 1.45-1.65 (4H, m), 1.90-2.10 (8H, m), 2.24 (6H,s), 2.48 (2H, t, J=6 Hz), 3.18 (2H, t, J=6 Hz), 3.40-3.60 (6H, m),3.60-3.75 (1H, m), 4.08-4.30 (2H, m), 5.30-5.40 (4H, m) MS (FAB): 743(M+H)⁺

Example of Synthesis-12 Synthesis of 2-dimethylaminoethylN-(2,3-dioleyloxypropyl)carbamate

In 2 ml of pyridine was dissolved 45 mg (0.5 mmol) of2-dimethylaminoethanol followed by addition of 97 mg (0.6 mmol) ofN,N'-carbonyldiimidazole and the mixture was stirred for 4 hours. Tothis solution was added 355 mg (0.6 mmol) of 2,3-dioleyloxypropylaminedropwise and the mixture was stirred for 24 hours. After completion ofthe reaction, the solvent was distilled off and the residue wasdissolved in methylene chloride, washed with saturated aqueous sodiumhydrogen carbonate, dried, and concentrated. The residue was purified bycolumn chromatography (silica gel/methylene chloride-methanol) toprovide 383 mg (100%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.87 (6H, t, J=6 Hz, CH₃ ×2), 1.12-1.44 (44H,m, CH₂ ×22), 1.46-1.64 (4H, m, OCH₂ CH₃ ×2), 1.88-2.12 (8H, m, CH═CHCH₂×4), 2.37 (6H, s, N(CH₃)₂), 2.54 (2H, t, J=6 Hz, NC H₂), 3.32-3.64 (9H,m, OCH₂ ×3, OCH and NHCH₂), 4.16 (2H, t, J=6 Hz, COOCH₂), 5.17 (1H,brs., NHCO), 5.26-5.46 (4H, m, CH═CH×2) MS (FAB): 707 (M+H)⁺

Example of Synthesis-13 Synthesis of2-O-(2-dimethylaminoethyl)carbamoyl-1,3-O-dioleylglycerol

To 2 ml of a solution of 150 mg (0.253 mmol) 1,3-O-dioleylglycerol inpyridine was added 82 mg (0.51 mmol) of N,N'-carbonyldiimidazole and themixture was stirred at ambient temperature for 5 hours. The solvent wasthen distilled off under reduced pressure and the residue was dissolvedin methylene chloride, washed with 5% sodium dihydrogen phosphate-water,dried, and concentrated. The residue was dissolved in 1.6 ml ofN,N-dimethylformamide and stirred together with 45 mg (0.51 mmol) ofN,N-dimethyl-ethylenediamine overnight. After completion of thereaction, the solvent was distilled off and the residue was diluted withwater and extracted with methylene chloride. The extract was washed withwater, dried, and concentrated. The residue was subjected to columnchromatography (silica gel/chloroform-methanol) to provide 179 mg (100%)of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (44H, brs.),1.50-1.65 (4H, m), 1.90-2.10 (8H, m), 2.20 (6H, s), 2.39 (2H, t, J=6Hz), 3.20-3.30 (2H, m), 3.34-3.55 (4H, m), 3.55-3.70 (4H, d, J=4 Hz),4.99 (1H, t, J=4 Hz), 5.25-5.46 (5H, m) MS (FAB): 707 (M+H)⁺

Example of Synthesis-14 Synthesis of 2-dimethylaminoethylN-(1,3-dioleyloxypropan-2-yl)carbamate

Using 1,3-dioleyloxy-2-propylamine, the procedure of Example ofSynthesis-12 was otherwise repeated to provide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (44H, brs.),1.50-1.60 (4H, m), 1.90-2.10 (8H, m), 2.28 (6H, s), 2.54 (2H, t, J=6Hz), 3.40-3.55 (8H, m), 3.80-3.90 (1H, m), 4.15 (2H, t, J=6 Hz),5.10-5.20 (1H, m), 5.20-5.45 (4H, m) MS (FAB): 707 (M+H)⁺

Example of Synthesis-15 Synthesis of3-O-(2-dimethylaminoethyl)carbamoyl-1,2-O-dioleoylglycerol

Using 1,2-O-dioleoylglycerol, the procedure of Example of Synthesis-1was otherwise repeated to provide the title compound. ¹ H-NMR (200 MHz,CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (40H, brs.), 1.50-1.70 (4H, m),1.90-2.10 (8H, m), 2.22 (6H, s), 2.24-2.40 (4H, m), 2.41 (2H, t, J=6Hz), 3.20-3.30 (2H, m), 4.10-4.15 (4H, m), 5.20-5.30 (2H, m), 5.30-5.45(4H, m) MS (FAB): 735 (M+H)⁺

Example of Synthesis-16 Synthesis of2-O-(2-dimethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol

Using 1,3-O-dioleoylglycerol, the procedure of Example of Synthesis-13was otherwise repeated to provide the title compound. ¹ H-NMR (200 MHz,CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.26 (40H, brs.), 1.50-1.70 (4H, m),1.90-2.10 (8H, m), 2.22 (6H, s), 2.32 (4H, t, J=8 Hz), 2.42 (2H, t, J=6Hz), 3.20-3.30 (2H, m), 4.12-4.25 (4H, m), 5.15 (1H, t, J=6 Hz),5.20-5.45 (5H, m) MS (FAB): 735 (M+H)⁺

Example of Synthesis-17 Synthesis of 2-dimethylaminoethylN-(2,3-dioleoyloxypropyl)carbamate

In 30 ml of anhydrous pyridine was dissolved 500 mg (5.61 mmol) of2-dimethylaminoethanol followed by addition of 1.91 g (11.8 mmol) ofN,N'-carbonyldiimidazole and the reaction was conducted at ambienttemperature for 5 hours. To this reaction mixture was added 197 mg (2.16mmol) of 3-amino-1,2-propanediol and the reaction was carried out atambient temperature overnight. The pyridine was then distilled off underreduced pressure and the resulting crude carbamate was redissolved inanhydrous pyridine. Then, under ice-cooling, 5.22 g (17.4 mmol) ofoleoyl chloride was added and the reaction was conducted at 50° C. for14 hours. The pyridine was then distilled off under reduced pressure andthe residue was dissolved in methylene chloride and washed withsaturated aqueous sodium hydrogen carbonate solution. The methylenechloride layer was dried over sodium sulfate and concentrated. Theresidue was subjected to column chromatography (silica gel/methylenechloride-methanol) to provide 250 mg (16%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.25 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.28 (6H, s), 2.30 (4H, t, J=8Hz), 2.57 (2H, t, J=6 Hz), 3.30-3.50 (2H, m), 4.06-4.30 (4H, m),5.04-5.15 (2H, m), 5.25-5.40 (4H, m) MS (FAB): 735 (M+H)⁺

Example of Synthesis-18 Synthesis of 2-dimethylaminoethylN-(1,3-dioleoyloxypropan-2-yl)carbamate

Using 2-amino-1,3-propanediol in lieu of 3-amino-1,2-propanediol, theprocedure of Example of Synthesis-17 was otherwise repeated to provide372 mg (2.2 mmol) of the title compound. ¹ H-NMR (200 MHz, CDCl₃) δ:0.87(6H, t, J=7 Hz), 1.20-1.40 (40H, m), 1.50-1.70 (4H, m), 1.90-2.10 (8H,m), 2.30 (6H, s), 2.32 (4H, t, J=8 Hz), 2.59 (2H, t, J=6 Hz), 4.00-4.25(7H, m), 5.10-5.20 (1H, m), 5.30-5.45 (4H, m) MS (FAB): 735 (M+H)⁺

Example of Synthesis-19 Synthesis of2-O-(2-piperdinoethyl)carbamoyl-1,3-O-dioleoylglycerol

Using 1,3-O-dioleoylglycerol and 1-(2-aminoethyl)piperidine, theprocedure of Example of Synthesis-13 was otherwise repeated to providethe title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (40H, brs.),1.44-1.54 (2H, m), 1.54-1.76 (8H, m), 1.90-2.10 (8H, m), 2.32 (4H, t,J=8 Hz), 2.39-2.56 (6H, m), 3.20-3.40 (2H, m), 4.12-4.40 (4H, m),5.08-5.24 (1H, m), 5.24-5.52 (5H, m) MS (FAB): 773 (M+H)⁺

Example of Synthesis-20 Synthesis of2-O-(2-diethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol

Using 1,3-O-dioleoylglycerol and N,N-diethylethylenediamine, theprocedure of Example of Synthesis-13 was otherwise repeated to providethe title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.02 (6H, t, 6 Hz),1.28 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.32 (4H, t, J=8Hz), 2.44-2.66 (6H, m), 3.16-3.32 (2H, m), 4.22-4.38 (4H, m), 5.08-5.22(1H, m), 5.26-5.52 (5H, m) MS (FAB): 763 (M+H)⁺

Example of Synthesis-21 Synthesis of2-O-(2-diisopropylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol

Using 1,3-O-dioleoylglycerol and N,N-diisopropylethylenediamine, theprocedure of Example of Synthesis-13 was otherwise repeated to providethe title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.00 (12H, t, 6 Hz),1.27 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.30 (4H, t, J=8Hz), 2.48-2.64 (2H, m), 2.88-3.20 (4H, m), 4.10-4.32 (4H, m), 5.06-5.28(2H, m), 5.30-5.42 (4H, m) MS (FAB): 791 (M+H)⁺

Example of Synthesis-22 Synthesis of2-O-(2-pyrrolidinoethyl)carbamoyl-1,3-O-dioleoylglycerol

Using 1,3-O-dioleoylglycerol and 1-(2-aminoethyl)pyrrolidine, theprocedure of Example of Synthesis-13 was otherwise repeated to providethe title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.27 (40H, brs.),1.50-1.70 (4H, m), 1.74-1.88 (4H, m), 1.90-2.10 (8H, m), 2.30 (4H, t,J=8 Hz), 2.44-2.70 (6H, m), 3.20-3.40 (2H, m), 4.20-4.42 (4H, m),5.08-5.22 (1H, m), 5.24-5.46 (5H, m) MS (FAB): 761 (M+H)⁺

Example of Synthesis-23 Synthesis of2-O-(2-morpholinoethyl)carbamoyl-1,3-O-dioleoylglycerol

Using 1,3-O-dioleoylglycerol and 4-(2-aminoethyl)morpholine, theprocedure of Example of Synthesis-13 was otherwise repeated to providethe title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.27 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.31 (4H, t, J=8 Hz), 2.40-2.54(6H, m), 3.20-3.40 (2H, m), 3.70 (4H, t, J=6 Hz), 4.12-4.38 (4H, m),5.08-5.20 (2H, m), 5.20-5.46 (4H, m) MS (FAB): 777 (M+H)⁺

Example of Synthesis-24 Synthesis of2-O-(3-diethylaminopropyl)carbamoyl-1,3-O-dioleoylglycerol

Using 1,3-O-dioleoylglycerol and 3-diethylaminopropylamine, theprocedure of Example of Synthesis-13 was otherwise repeated to providethe title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.03 (6H, t, 6 Hz),1.28 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.30 (4H, t, J=8Hz), 2.46-2.58 (6H, m), 3.20-3.32 (2H, m), 4.10-4.34 (4H, m), 5.10-5.20(1H, m), 5.30-5.42 (4H, m), 6.18-6.30 (1H, brs.) MS (FAB): 777 (M+H)⁺

Example of Synthesis-25 Synthesis of 2-O-2-(N-methyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

In 10 ml of chloroform was dissolved 173 mg (0.224 mmol) of1,3-O-dioleoyl-2-O-(2-bromoethyl)carbamoylglycerol followed by additionof 543 mg (7.228 mmol) of 2-(methylamino)ethanol and 27 mg (0.209 mmol)of diisopropylethylamine and the mixture was refluxed at 80° C.overnight. The reaction mixture was then washed with 5% sodiumdihydrogen phosphate-H₂ O, dried, and concentrated. The residue wassubjected to column chromatography (silica gel/methylenechloride-methanol) to provide 128 mg (74.3%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.86 (6H, t, J=6 Hz), 1.27 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.26-2.38 (7H, m), 2.50-2.70 (4H,m), 3.20-3.40 (2H, m), 3.61 (4H, t, J=6 Hz), 4.20-4.44 (4H, m),5.06-5.20 (2H, m), 5.30-5.45 (4H, m) MS (FAB): 765 (M+H)⁺

Example of Synthesis-26 Synthesis of 2-O-2-(N-ethyl-N-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

Using 2-(ethylamino)ethanol, the procedure of Example of Synthesis-25was otherwise repeated to provide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.03 (3H, t, 6 Hz),1.28 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.32 (4H, t, J=8Hz), 2.54-2.68 (6H, m), 3.20-3.30 (2H, m), 3.56 (2H, t, J=6 Hz),4.12-4.34 (4H, m), 5.06-5.20 (2H, m), 5.30-5.44 (4H, m) MS (FAB): 779(M+H)⁺

Example of Synthesis-27 Synthesis of 2-O-2-(N,N-di-(2-hydroxyethyl)amino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

Using diethanolamine, the procedure of Example of Synthesis-25 wasotherwise repeated to provide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.28 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.32 (4H, t, J=8 Hz), 2.60-2.70(6H, m), 3.20-3.30 (2H, m), 3.60 (4H, t, J=6 Hz), 4.12-4.40 (4H, m),5.08-5.20 (1H, m), 5.30-5.42 (4H, m), 5.60-5.70 (1H, brs.) MS (FAB): 795(M+H)⁺

Example of Synthesis-28 Synthesis of 2-O-2-(N-methyl-N-n-butylamino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

Using N-methylbutylamine, the procedure of Example of Synthesis-25 wasotherwise repeated to provide the title compound. ¹ H-NMR (200 MHz,CDCl₃) δ:0.82-0.96 (9H, m), 1.10-1.50 (42H, m), 1.50-1.75 (6H, m),1.90-2.10 (8H, m), 2.19 (3H, s), 2.26-2.40 (6H, m), 2.46 (2H, m),3.20-3.30 (2H, m), 4.10-4.30 (4H, m), 5.08-5.20 (1H, m), 5.25-5.40 (4H,m) MS (FAB): 777 (M+H)⁺

Example of Synthesis-29 Synthesis of 2-O-2-(4-(2-hydroxyethyl)piperazino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

Using 1-(2-hydroxyethyl)piperazine, the procedure of Example ofSynthesis-25 was otherwise repeated to provide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (3H, t, J=6 Hz), 1.28 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.32 (4H, t, J=8 Hz), 2.40-2.60(12H, m), 3.18-3.32 (2H, m), 3.62 (2H, t, J=6 Hz), 4.12-4.32 (4H, m),5.08-5.24 (2H, m), 5.30-5.40 (4H, m) MS (FAB): 820 (M+H)⁺

Example of Synthesis-30 Synthesis of 2-O-2-(N,N,N',N'-tetramethylguanidino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

Using N,N,N',N'-teramethylguanidine, the procedure of Example ofSynthesis-25 was otherwise repeated to provide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (3H, t, J=6 Hz), 1.27 (40H, brs.),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.30 (4H, t, J=8 Hz), 2.96 (3H,s), 3.10 (3H, s), 3.35-3.40 (2H, m), 3.60-3.70 (2H, m), 4.04-4.34 (4H,m), 4.98-5.08 (1H, m), 5.30-5.40 (4H, m), 6.30-6.40 (1H, m) MS (FAB):805 (M+H)⁺

Example of Synthesis-31 Synthesis of 2-O-2-(N-(2-diethylamino)ethyl-N-methylamino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

Using N,N-diethyl-N'-methylethylenediamine, the procedure of Example ofSynthesis-25 was otherwise repeated to provide the title compound. ¹H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.04 (6H, t, J=6 Hz),1.26 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.26-2.36 (7H,m), 2.44-2.64 (10H, m), 3.15-3.25 (2H, m), 4.16-4.26 (4H, m), 5.08-5.18(1H, m), 5.30-5.40 (4H, m), 6.46-6.60 (1H, brs.) MS (FAB): 820 (M+H)⁺

Example of Synthesis-32 Synthesis of 2-O-2-(4-ethylpiperazino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

Using 1-ethylpiperazine, the procedure of Example of Synthesis-25 wasotherwise repeated to provide the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.10 (3H, t, J=6 Hz),1.26 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.32 (4H, t, J=8Hz), 2.38-2.60 (12H, m), 3.22-3.34 (2H, m), 4.12-4.34 (4H, m), 5.10-5.30(2H, m), 5.30-5.42 (4H, m) MS (FAB): 802 (M+H)⁺

Example of Synthesis-33 Synthesis of 2-O-2-(N-ethyl-N-methylamino)ethyl!carbamoyl-1,3-O-dioleoylglycerol

In 3 ml of chloroform was dissolved 131 mg (0.170 mmol) of1,3-O-dioleoyl-2-O-(2-bromoethyl) carbamoylglycerol followed by additionof 470 mg (7.951 mmol) of N-ethylmethylamine and the reaction wasconducted in a sealed tube at 80° C. overnight. This reaction mixturewas then washed with 5% sodium dihydrogen phosphate-H₂ O, dried andconcentrated. The residue was subjected to column chromatography (silicagel/methylene chloride-methanol) to provide 104 mg (81.5%) of the titlecompound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.04 (3H, t, J=6 Hz),1.26 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.20 (3H, s),2.32 (4H, t, J=8 Hz), 2.38-2.52 (4H, m), 3.20-3.30 (2H, m), 4.12-4.32(4H, m), 5.10-5.20 (1H, m), 5.25-5.42 (5H, m) MS (FAB): 749 (M+H)⁺

Example of Synthesis-34 Synthesis of2-O-(2-diethylaminoethyl)carbamoyl-1,3-O-dipalmitoylglycerol

Using 1,3-O-dipalmitoylglycerol and N,N-diethylethylenediamine, theprocedure of Example of Synthesis-13 was otherwise repeated to providethe title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.87 (6H, t, J=6 Hz), 1.00 (6H, t, J=6 Hz),1.25 (48H, brs.), 1.50-1.70 (4H, m), 2.30 (4H, t, J=8 Hz), 2.46-2.60(6H, m), 3.10-3.30 (2H, m), 4.12-4.32 (4H, m), 5.10-5.20 (1H, m),5.20-5.35 (1H, m) MS (FAB): 711 (M+H)⁺

Example of Synthesis-35 Synthesis of 2-diethylaminoethylN-(1,3-dioleoyloxypropan-2-yl)carbamate

In methylene chloride was dissolved 470 mg (4 mmol) of2-diethylaminoethanol. After addition of 633 mg (8 mmol) of pyridine,690 mg (4.4 mmol) of phenyl chloroformate was further added underice-cooling and the reaction was conducted at ambient temperature for 2hours. After completion of the reaction, the solvent was distilled offand the residue was transferred into ethyl acetate-1% aqueous sodiumhydrogen carbonate solution. The ethyl acetate layer was separated,dried over sodium sulfate, and concentrated under reduced pressure togive 705 mg (74%) of crude carbonate compound. This crude carbonate wasdissolved in anhydrous pyridine followed by addition of 134 mg (1.47mmol) of 2-amino-1,3-propanediol and the reaction was carried out at 80°C. overnight. Then, 973 mg (3.2 mmol) of oleoyl chloride was added andthe reaction was further conducted at ambient temperature for 24 hours.After completion of the reaction, the solvent was distilled off underreduced pressure and the residue was transferred into methylenechloride-saturated aqueous sodium hydrogen carbonate solution and driedover sodium sulfate. The solvent was then removed under reduced pressureand the residue was subjected to column chromatography (silicagel/methylene chloride-methanol) to provide 250 mg (22%) of the titlecompound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.87 (6H, t, J=6 Hz), 1.04 (6H, t, J=6 Hz),1.28 (40H, brs.), 1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.30 (4H, t, J=8Hz), 2.50-2.70 (6H, m), 4.00-4.30 (7H, m), 5.05-5.20 (1H, m), 5.25-5.45(4H, m) MS (FAB): 763 (M+H)⁺

Example of Synthesis-36 Synthesis of2-O-(3-diethylaminopropionyl)-1,3-O-dioleoylglycerol

In a solvent mixture of 3 ml N,N-dimethylformamide and 6 ml methylenechloride was dissolved 172 mg (0.277 mmol) of 1,3-dioleoylglycerolfollowed by addition of 101 mg (0.556 mmol) of N,N-diethyl-β-alanine(hydrochloride), 114 mg (0.553 mmol) of N,N-dicyclohexylcarbodiimide,and 7 mg (0.057 mmol) of 4-dimethylaminopyridine and the mixture wasstirred overnight. This reaction mixture was then filtered and thefiltrate was concentrated under reduced pressure. The residue wasdissolved in methylene chloride and washed with water. The washedsolution was dried and concentrated and the residue was subjected tocolumn chromatography (silica gel/methylene chloride-methanol) toprovide 129 mg (62%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 1.20-1.40 (46H, m),1.50-1.70 (4H, m), 1.90-2.10 (8H, m), 2.32 (4H, t, J=8 Hz), 2.76-2.84(6H, m), 3.04-3.14 (2H, m), 4.08-4.42 (4H, m), 5.18-5.30 (1H, m),5.30-5.44 (4H, m) MS (FAB): 748 (M+H)⁺

Example of Synthesis-37 Synthesis ofO-(2-dimethylaminoethyl),O'-(1,3-dioleoyloxypropyl)methylphosphonate

To 310 mg (0.50 mmol) of 1,3-dioleoylglycerol dried by azeotropicdistillation with pyridine was added 9.1 ml (1 mmol) of 0.11M methylbis-O,O-(1-benzotriazolyl)phosphonate-dioxane and the reaction wasconducted at ambient temperature for 3 hours. To this reaction mixturewere added 446 mg (5 mmol) of 2-dimethylaminoethanol and 411 mg (5 mmol)of 1-methylimidazole and the reaction was further conducted at ambienttemperature overnight. The reaction mixture was then treated withmethylene chloride-5% sodium dihydrogen phosphate solution and themethylene chloride layer was dried over sodium sulfate and concentratedunder educed pressure. The residue was subjected to columnchromatography (silica gel/methylene chloride-methanol) to provide 272mg (59%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.86 (6H, t, J=6 Hz), 1.25 (40H, brs), 1.54(3H, d, J=20 Hz), 1.50-1.70 (4H, m), 2.32 (4H, t, J=8 Hz), 2.35 (6H, s),2.68 (2H, t, J=6 Hz), 4.05-4.25 (4H, m), 4.25-4.35 (2H, m), 4.70-4.90(1H, m), 5.25-5.40 (4H, m) MS (FAB): 770 (M+H)⁺

Example of Synthesis-38 Synthesis ofO-(2-aminoethyl)-O'-(1,3-dioleoyloxypropyl)methylphosphonate

Using t-butyl N-(2-hydroxyethyl)carbamate in lieu of2-dimethylaminoethanol, the procedure of Example of Synthesis-37 wasotherwise repeated and the resulting compound was treated withtrifluoroacetic acid/methylene chloride (1:2) to provide the titlecompound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.88 (6H, t, J=6 Hz), 2.25 (40H, brs),1.50-1.90 (7H, m), 1.90-2.10 (8H, m), 2.34 (4H, t, J=8 Hz), 3.30-3.40(2H, s), 4.10-4.50 (6H, m), 4.75-4.90 (1H, m), 5.30-5.40 (4H, m) MS(FAB): 742 (M+H)⁺

Example of Synthesis-39 Synthesis of O-(2-diethylaminoethyl)-O'-(1,3-dioleoyloxypropyl)methylphosphonate

Using 2-diethylaminoethanol, the procedure of Example of Synthesis-37was otherwise repeated to provide 166 mg (70.7%) of the title compound.

¹ H-NMR (200 MHz, CDCl₃) δ:0.87 (6H, t, J=6 Hz), 1.01 (6H, t, J=6 Hz),1.26 (40H, brs), 1.48-1.70 (7H, m), 1.90-2.10 (8H, m), 2.32 (4H, t, J=8Hz), 2.57 (4H, q, J=6 Hz), 2.80 (2H, t, J=6 Hz), 3.90-4.40 (6H, m),4.70-4.90 (1H, m), 5.30-5.42 (4H, m) MS (FAB): 798 (M+H)⁺

Production Example 1

In 200 μl of chloroform, in a vial, were dissolved 5 mg of the compoundaccording to Example of Synthesis-4 and 5 mg of egg yolkphosphatidylethanolamine. Then, nitrogen gas was blasted against thesolution.to remove the chloroform, leaving a thin film on the internalwall of the vial. The vial was then allowed to stand overnight underreduced pressure and after addition of 2 ml of sterile distilled water,was agitated in a vortex mixer to exfoliate the thin film. After purgingwith nitrogen gas, the vial was hermetically stoppered and allowed tostand at 4° C. for 3 hours. Then, sonication was carried out for 10minutes with a bath sonicator to provide a lipid device.

Production Example 2

Using the compound according to Example of Synthesis-1, a lipid devicewas prepared in otherwise the same manner as Production Example 1.

Production Example 3

Using the compound according to Example of Synthesis-7, a lipid devicewas prepared in otherwise the same manner as Production Example 1.

Production Example 4

In 200 l of chloroform, in a vial, were dissolved 5 mg of the compoundaccording to Example of Synthesis-20 and 5 mg of egg yolkphosphatidylcholine. Then, nitrogen gas was blasted against the solutionto remove the chloroform, leaving a thin film on the internal wall ofthe vial. The vial was then allowed to stand overnight under reducedpressure and after addition of 2 ml of sterile distilled water, wasagitated in a vortex mixer to exfoliate the thin film. After purgingwith nitrogen gas, the vial was hermetically stoppered and allowed tostand at 4° C. for 3 hours. Then, sonication was carried out for 10minutes with a bath sonicator to provide a lipid device.

Production Example 5

Using the compound according to Example of Synthesis-1, a lipid devicewas prepared in otherwise the same manner as Production Example 4.

Production Example 6

Using the compound according to Example of Synthesis-4, a lipid devicewas prepared in otherwise the same manner as Production Example 4.

Production Example 7

Using the compound according to Example of Synthesis-7, a lipid devicewas prepared in otherwise the same manner as Production Example 4.

Example 1 An alternate poly(adenylic acid-uridylic acid)-containinginjectable composition

To 60 μl of the lipid device according to Production Example 4 was added0.9 ml of physiological saline solution, followed by addition of 0.1 mlof a 100 μg/ml saline solution of an alternate poly(adenylicacid-uridylic acid) Poly(rA-rU)• Poly(rA-rU), S20, w=4.70, manufacturedby Pharmacia; the same applies hereinafter!, and the mixture was stirredto provide an injectable composition. Similar injectable compositionswere prepared using the lipid devices according to Production Examples1-3 and 5-7.

Example 2 An alternate poly(adenylic acid-uridylic acid)-containinginjectable composition

To 300 μl of Lipofectin (trademark, Bethesda Laboratories) was added 0.9ml of physiological saline, followed by addition of 0.1 ml of a 100μg/ml saline solution of the alternate polymer and the mixture wasstirred to provide an injectable composition.

Test Example 1 HeLaS3 cell growth inhibitory action (in vitro)

A 96-well plate was seeded with HeLaS3 cells at a cell density of 10⁴cells/well (90 μl). On the following day, 10 μl of the 30 μg/ml lipiddevice containing a varying concentration of alternate poly(adenylicacid-uridylic acid) was added to each well. As to the compositioncontaining Genetransfer (trademark, Wako Pure Chemical Industries), 1 mlof 10 mM phosphate buffer (supplemented with 0.9% sodium chloride)containing 20 μg of dissolved alternate poly (adenylic acid-uridylicacid) was added into each vial of Genetransfer and this was used asdiluted to predetermined concentrations of alternate poly(adenylicacid-uridylic acid). The plate was incubated for 72 hours after additionand, then, 10 μl per well of a 5 mg/ml solution of MTT(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide was added.After 2-4 hours, the reaction was stopped by adding isopropylalcohol/0.04N hydrochloric acid mixture. After each well was agitated,the absorbance at 540 nm was measured with a plate reader (manufacturedby Corona) and the percentage HeLaS3 cell growth inhibition (%) wascalculated. This calculation was performed according to the equationgiven below. The cell growth inhibition rate of the single-strandednucleic acid copolymer in the absence of the lipid device was taken ascontrol. ##EQU1##

The results are shown in Table 1.

                  TABLE 1                                                         ______________________________________                                        Concn. of                                                                              % Inhibition                                                         poly(A-U)                     Exp. of Syn.-1 +                                (ng/ml)  Control    genetransfer                                                                            EPE                                             ______________________________________                                        0        0          0         0                                               0.1      0          15        1                                               1        0          0         36                                              10       0          32        91                                              100      0          82        100                                             ______________________________________                                        Concn. of  % Inhibition                                                       poly(A-U)  Exp. of Syn.-4     Exp. of Syn.-20                                 (ng/ml)    +EPE    +ELC       +EPE  +ELC                                      ______________________________________                                        0          0       0          0     0                                         0.1        0       0          25    25                                        1          0       0          62    62                                        10         61      65         89    89                                        100        100     91         100   100                                       ______________________________________                                         genetransfer: trademark, Wako Pure Chemical Industries                        EPE; egg yolk phosphatidylethanolamine                                        ELC; egg yolk phosphatidylcholine                                             Exp. of Syn. = Example of Synthesis                                      

It is apparent from Table 1 that when applied together with the lipiddevice, the single-stranded nucleic acid copolymer alternatepoly(adenylic acid-uridylic acid)! which is not effective when usedalone shows cell growth inhibitory activity.

Test Example 2 Induction of β-interferon from HeLaS3 cells

A 96-well plate was seeded with 10⁴ cells/well of HeLaS3 cells (90 μl)and on the following day 10 μl of the 30 μg/ml lipid device containing avarying concentration of alternate poly(adenylic acid-uridylic acid) wasadded to each well. The plate was incubated for 24 hours after additionand the β-interferon in the cell culture was determined using an ELISAkit (for assay of β-interferon, manufactured by Toray Industries, Inc.).The results are shown in Table 2.

                  TABLE 2                                                         ______________________________________                                        Concn. of                                                                     poly (A-U)    IU/ml                                                           (ng/ml)       Control Exp. of Syn.-4 + EPE                                    ______________________________________                                        0             0       0                                                       0.1           0       10                                                      1             0       34                                                      10            0       32                                                      100           0       148                                                     ______________________________________                                         EPE; egg yolk phosphatidylethanolamine                                        Exp. of Syn. 4 = Example of Synthesis4                                   

What is claimed is:
 1. A composition comDrisinq a lipid device and asingle-stranded nucleic acid copolymer, wherein said lipid device is amixture of a phospholipid and a compound of the following generalformula ##STR17## wherein R¹ and R² are not the same and each representsOY or --A--(CH₂)n-E. n represents a whole number of 0-4. E representspyrrolidino, piperidino, substituted or unsubstituted piperazino,morpholino, substituted or unsubstituted guanidino, or ##STR18## (R³ andR⁴ are the same or different and each represents hydrogen, lower(C₁ -₄)alkyl, hydroxy-lower(C₁ -₄) alkyl, or mono- or di-(lower)alkylaminoalkyl(C₂₋₆)). A represents the following 1, 2, 3, 4, 5, 6, or7 ##STR19## R and Y are the same or different and each represents asaturated or unsaturated aliphatic hydrocarbon group of 10-30 carbonatoms or a saturated or unsaturated fatty acid residue of 10-30 carbonatoms.
 2. A composition comprisinq a lipid and a single-stranded nucleicacid copolymer, wherein said lipid is a mixture of a phospholipid and acompound of the following general formula ##STR20## wherein R¹¹ and R²¹are the same or different and each represents oleyl or oleoyl. Drepresents --CH₂ -- or --NH--. R³⁰ and R⁴⁰ are the same or different andeach represents methyl or ethyl.
 3. A composition according to claim 1,wherein said lipid is a mixture of a phospholipid and a compoundselected from the group consisting of3-O-(4-dimethylaminobutanoyl)-1,2-O-dioleylglycerol,3-O-(2-dimethylaminoethyl)carbamoyl-1,2-O-dioleylglycerol, and2-O-(2-diethylaminoethyl)carbamoyl-1,3-O-dioleoylglycerol.
 4. Acomposition according to claim 1, wherein said phospholipid is selectedfrom the group consisting of phosphatidylethanolamine andphosphatidylcholine.
 5. A composition accordinq to claim 1, wherein saidsingle-stranded nucleic acid copolymer is poly(adenylic acid-uridylicacid).
 6. A composition according to claim 2, wherein said phospholipidis selected from the group consisting of phosphatidylethanolamine andphosphtidylcholine.
 7. A composition according to claim 3, wherein saidphospholipid is selected from the group consisting ofphosphatidylethanolamine and phosphatidylcholine.
 8. A compositionaccording to claim 2, wherein said single-stranded nucleic acidcopolymer is poly(adenylic acid-uridylic acid).
 9. A compositionaccording to claim 3, wherein said single-stranded nucleic acidcopolymer is poly(adenylic acid-uridylic acid).
 10. A compositionaccording to claim 4, wherein said single-stranded nucleic acidcopolymer is poly(adenylic acid-uridylic acid).